RIP-Tag2 mouse model as a Paradigm for Target Search in NETs Oriol Casanovas, Ph.D. Tumor Angiogenesis Group INSTITUT CATALÀ d ONCOLOGIA IDIBELL Barcelona (SPAIN)
Therapeutic Targeting of the Tumor Stroma Joyce J, Cancer Cell 2005
Mechanisms of Tumor Angiogenesis
First Preclinical Models of NET NET Cell culture Injection into mice BON cells (CJ Thompson 1991) GOT1 cells (Kölby & Nisson 2000) Develop Tumors
First Preclinical Models of NET BON, GOT1 cells injected subcutaneously into Nude Mice: GOOD PRECLINICAL MODEL OF CARCINOID TUMORS! CAVEATS: - Only models of Carcinoids are available - Artificiality of cell culture growth - Subcutaneous growth of tumors is a limitation: - Different tumor growth kinetics - Lack of proper tumor stroma Limitations of their use for target discovery!
Another model of NET: Transgenic Mouse Model of Insulinoma RIP-Tag2 Transgene: Rat Insulin Promoter SV40 Large + Small T antigen prb p53 Insulinoma Tumors Acinar Tumor Hanahan D. Nature 1985
RIP-Tag2: Multi-Stage Development of NET Normal (onc+) Hyperplastic/ Dysplastic Angiogenic Tumor <5 wks 100% 5-77 wks ~50% 7-12 wks ~10% 12-14 14 wks 2-4% Hanahan D. Nature 1985
RIP-Tag2: Distinct Neoplastic Lesions Distinct neoplastic lesions in RIP-Tag pancreas: Hanahan D. Nature 1985
RIP-Tag2: Quantitation of Tumorigenesis RIP1-Tag2 Pancreas Normal Pancreas 10wk 12wk Normal Islets Angiogenic Islets Tumors
Stage-Specific Experimental Therapeutic Trial Designs Prevention 5-8 wks of age Intervention 10-13.5 wks of age Regression 12-16 wks of age Hyperplastic/ Dysplastic Angiogenic Tumor Prevention trial: Can angiogenic switching be prevented? Intervention trial: Can tumor formation be prevented or slowed? Regression trial: Can tumor growth be stabilized or regressed d and lifespan be extended?
Antiangiogenic VEGFR inhibitors are effective in the RIP-Tag2 model DC101: Anti-VEGFR2 Ab specifically blocks activation of mvegfr2 Casanovas et al. Cancer Cell 2005
Anti-VEGFR2 Specific Blocking Antibody is fairy effective DC101: Anti-VEGFR2 Ab specifically blocks activation of mvegfr2. 35 Angiogenic Switch 180 Tumor formation 160 Tumor Growth Number of Angiogenic "red" Islets 30 25 20 15 10 5 * Tumor Burden (mm3) 160 140 120 100 80 60 40 20 * Tumor Burden (mm3) 140 120 100 80 60 40 20 * 0 Ctrl Anti-R2 0 Ctrl Anti-R2 0 Ctrl Ctrl Anti-R2 t=0 10 days Casanovas et al. Cancer Cell 2005
Perivascular cells, PERICYTES, in tumor vasculature PDGFR β / CD31 PDGFR β / lectin
The concept of multi-targeted inhibition of tumor angiogenesis Bergers, et al. (2003). Benefits of targeting both pericytes and endothelial cells in tumor vasculature with kinase inhibitors. J.C.I., 111: 1287-95
The concept of multi-targeted inhibition of tumor angiogenesis Bergers, et al. (2003). Benefits of targeting both pericytes and endothelial cells in tumor vasculature with kinase inhibitors. J.C.I., 111: 1287-95
Antiangiogenic VEGFR small molecule inhibitors Sunitinib, Sorafenib, Pazopanib, PTK787 Favine et al. NRDD 2007
Vascular Regression Kinetics on Sunitinib Treatment Antiangiogenic effects in the RIP-Tag2 islet cell cancer tumor model: Control Sunitinib 4d Sunitinib 7d Sunitinib 14d. Sunitinib 28d. Pericyte EC association After Sunitinib Treatment: Sunitinib 7d Sunitinib 14d Sunitinib 28d McDonald Lab Virginia Yao UCSF
Anti-Angiogenic (Sunitinib) therapies for Cancer Sunitinib treated Transgenic multistage mouse model of cancer: RIP-Tag2 (D.Hanahan) Resistance? Untreated, 14.5 w Sunitinib, 18 w
Anti-Angiogenesis: a non-resistant therapeutic approach? Chemotherapy AntiAngiogenesis Target: Tumor Cell Genetic Instability High adaptation Resistance Target: Endothelial Cell Genetically Stable Less Adaptation No Resistance?
DC101 anti-vegfr2 Blocking Antibodies in RIP-Tag2 Tumors DC101: Anti-VEGFR2 Ab specifically blocks activation of mvegfr2. 160 140 Tumor Burden (mm3) 120 100 80 60 * RESISTANCE TO ANTI-ANGIOGENESIS 40 20 0 Control Control Anti-VEGFR2 Anti-VEGFR2 t=0 10 days 4 weeks tt 12 wks of age Casanovas et al. Cancer Cell 2005
Test FGF family in the Tumor Regrowth Phase t=0 10 days 4 wks Stasis Phase Regrowth Phase Anti-VEGFR2 blocking antibody FGF TRAP Adenovirus-sFGFR2iiib 160 p=0.002 140 Tumor Burden (mm3) 120 100 80 60 40 20 0 Ctrl t=0 Ctrl 10 d Anti-R2 10 d Anti-R2 4 wks Anti-R2 FGF trap 4 wks
Tumor Responses to Anti-Angiogenesis Blocking of VEGFR2 with DC101 mab in the RIP-Tag2 model: VEGF Anti-R2 No Angiogenesis Hypoxia Anti-R2 VEGF other Factors Reactivation of Angiogenesis RESPONSE PHASE with decreased angiogenesis (v. density & permeability) and inhibition of tumor growth (tumor stasis ). RESISTANCE PHASE with reestablishment of the vasculature and tumor re-growth, that implicates FGFs and other pro-angiogenic factors. Casanovas et al. Cancer Cell 2005
Resistance to Anti-Angiogenesis therapies for Cancer P.Carmeliet Insight Review, Nature Dec. 2005
Tunmor Adaptation to Anti-Angiogeneic therapies ANTIANGIOGENIC THERAPY (inh. VEGF/VEGFR2) Vascularized Tumor with sufficient nutrients and oxygen supply Stop neovascularization, Vessel Pruning Decreased MVD, blood perfusion, Hypoxia anti-tumor effect. Tumor Reaction : - Adaptation - Hypoxia tolerance - Microenvironment adaptation RESISTANCE!!!
Responses to anti-vegfr2: Increased Invasive Phenotype Transgenic multistage mouse model of cancer: RIP-Tag2 (D.Hanahan) Antiangiogenic therapy: DC101 anti-mvegfr2 Blocking Antibody (1 mg/mouse twice a week IP) Control (end-stage) Anti-VEGFR2 1 week Anti-VEGFR2 4 weeks H&E 150 µm 150 µm 150 µm Ac Ac Ac Anti-T antigen T 50 µm T 50 µm 50 µm T Pàez-Ribes et al. Cancer Cell 2009
Adaptive Responses to anti-vegfr2: Increased Dissemination & Mets Pàez-Ribes et al. Cancer Cell 2009
Enhanced Metastasis in Intermitent and Continuous Sunitinib treatment! Sunitinib 40 mg/kg daily t=0 5 wks Anti-CD31 Control Sunitinib 5 weeks Invasion Incidence (%) 60 50 40 30 20 * ** Vehicle Sunitinib 10 Anti-T antigen Anti-T antigen 0 Incidence of metastasis (%) 50 40 30 20 10 0 Non invasive tumors Liver metastasis * Microinvasive tumors Highly invasive tumors LN metastasis Control Sunitinib Liz Allen & Douglas Hanahan (UCSF)
Conclusions and Perspectives In several animal models: -PNET tg -GBM orthotopic -BrCa human orthotopic -RCC human orthotopic -PanCa human orthotopic In some patients? ONGOING at ICO-IDIBELL: Which patients suffer this effects?
Acknowledgements External Collaborators Marta Páez* Mar Martínez Lozano Gabriela Jiménez Lidia Moserle Francesc Viñals Mariona Graupera Gabriel Capellà Univ. California San Francisco (US) Douglas Hanahan Gabriele Bergers Karolinska Institutet (Sue) Kristian Pietras Ins. Richerca e Cura del Cancro (IT) Enrico Giraudo Clinical Collaborators Ramon Salazar Àlex Teulé Margarita García